Size-Selective Nucleic Acid Enrichment via Polymer Elution
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Solution Overview
Problem
Current size-selective DNA enrichment methods for next-generation sequencing are time-consuming and costly, requiring multiple rounds of adapter removal steps using poly(alkylene oxide) polymers, which are inefficient and labor-intensive.
Innovation Solution
A method utilizing poly(alkylene oxide) polymers to bind target nucleic acid molecules of a specific size to a solid phase, followed by a size-selective elution step with a reagent composition containing a poly(alkylene oxide) polymer and a salt, allowing for the efficient removal of non-target nucleic acid molecules, thereby simplifying the process and reducing material and equipment requirements.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If multiple rounds of adapter removal steps using poly(alkylene oxide) polymers are performed, then adapter removal efficiency is improved, but processing time and operational complexity increase
Solution Approach 1:
The patent applies preliminary action by performing a size-selective elution step before the final elution of target DNA. This intermediate step removes non-target nucleic acids (adapter dimers and monomers) that would otherwise require multiple removal rounds, thereby reducing total processing time while maintaining adapter removal efficiency
Solution Approach 2:
The patent extracts non-target nucleic acid molecules (adapter dimers and monomers) from the sample during the size-selective elution step. By removing these contaminants early in the process, the need for multiple subsequent adapter removal steps is eliminated, reducing both time and operational complexity
2Reliability
If multiple rounds of adapter removal steps are performed, then adapter removal efficiency is improved, but material consumption and cost increase
Solution Approach 1:
The size-selective elution step is performed as a preliminary action before final DNA elution, removing non-target molecules in a single step. This eliminates the need for multiple rounds of adapter removal, thereby reducing consumption of poly(alkylene oxide) polymers, salts, and other reagents
Solution Approach 2:
The patent changes parameters by using a size-selective elution condition with specific poly(alkylene oxide) polymer concentration and salt type that preferentially elutes non-target molecules while retaining target DNA. This parameter optimization allows single-step contamination removal, reducing material consumption
3Reliability
If size-selective elution with poly(alkylene oxide) polymer and salt is performed, then non-target nucleic acid removal is improved, but process complexity increases
Solution Approach 1:
The patent merges the size-selective elution step with the existing adapter removal workflow, combining contamination removal with the standard protocol. This integration adds only one intermediate step rather than creating a separate complex process, maintaining simplicity while improving non-target molecule removal
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
This approach significantly reduces the time and cost of DNA enrichment, achieving equivalent adapter removal efficiency with fewer materials and lab equipment, while enabling easier automation and improved library yields.
Implementation Method 1
binding target nucleic acid molecules to a solid phase... wherein the bound nucleic acid molecules comprise target nucleic acid molecules
Implementation Method 2
contacting the solid phase with the bound nucleic acid molecules at least once with a reagent composition comprising a poly(alkylene oxide) polymer and a salt to selectively elute non-target nucleic acid molecules
Data Source
AI summary
The present invention provides a poly(alkylene oxide) polymer based size selective nucleic acid enrichment method for enriching target nucleic acid molecules from a nucleic acid containing sample which comprises target nucleic acid molecules and non-target nucleic acid molecules, wherein the target nucleic acid molecules are longer than the non-target nucleic acid molecules, the method comprising (a) preparing a binding mixture comprising—the nucleic acid containing sample, —a poly(alkylene oxide) polymer and—a salt and binding nucleic acid molecules to a solid phase which comprises a functional group, preferably carboxylated magnetic particles, wherein the bound nucleic acid molecules comprise target nucleic acid molecules; (b) preferably separating the solid phase with the bound nucleic acid molecules from the remaining sample; (c) contacting the solid phase with the bound nucleic acid molecules at least once with a reagent composition comprising a poly(alkylene oxide) polymer and a salt to selectively elute non-target nucleic acid molecules, wherein the concentration (w/v) of the poly(alkylene oxide) polymer in the reagent composition of step (c) is lower than the concentration (w/v) of the poly(alkylene oxide) polymer in the binding mixture of step (a); (d) optionally washing the bound target nucleic acid molecules; and (e) eluting the bound target nucleic acid molecules from the solid phase. Said method allows the size selective purification of target DNA molecules and is particularly suitable for sequencing applications. It is more time- and cost efficient than prior art methods and provides excellent purification results. Moreover, further methods and kits are provided.